Cell-cell interactions are essential to regulating the behavior and function of cells and tissues. Cell micro-patterning has become a very useful technique in cell biology, allowing precise control over the spatial organization of cell populations in vitro. This has enabled new types of experiments to be performed and unique insights into tissue biology. However, these techniques are typically developed in engineering laboratories, and high-quality patterning tools are not easily accessible to users with conventional training in cell biology.
The patterning of a sharp interface between two different cell populations, with direct cell-cell contact between the two populations, is useful for a number of different biological studies. A variety of tissue systems exhibit phenotypic differences when two different cell populations are mixed and allowed to interact. These include liver hepatocytes and non-parenchymal liver cells, endothelial and smooth muscle vascular cells, neurons and glial cells, neurons and meningeal cells, stem cells and feeder layers, and tumor and host stromal cells. A sharp patterned interface between two populations allows investigation of the role of direct cell-cell contact, gradients in cell signaling, migration and invasion between populations, morphogenesis, scarring and fibrosis, and other forms of cell-cell crosstalk.
There are two typical methods of patterning cell populations. The first requires sequential seeding, wherein one population of cells is patterned first, and later a second population of cells is added to fill in the unoccupied regions. Patterning of the first population is often accomplished by micro-contact printing of an adhesive protein, the use of a removable stencil, or microfluidic channels. See reference [1]. The challenge with this approach is that some cells from the second population attach in regions occupied by the first population, resulting in cross-contamination.
The second approach requires cell migration following removal of a barrier. The two cell populations are seeded simultaneously in two separate regions, with a removable barrier in between. After cell attachment, the barrier is removed and the cells can migrate towards each other to form a contact interface. The challenge with this approach is that the width of the barrier is often hundreds of micrometers. Thus, the cells have to travel quite a distance before the two populations interface. By the time this gap is closed (which may take up to 48 hours), the interface may be ragged and not very sharp.
To address the aforementioned challenges, another class of devices consists of discrete plates that are first seeded with different cell types and then moved together to form a sharp cell-cell interface. This approach also allows cells to grow to confluence and reach a quiescent state prior to the initiation of co-culture. In addition, the cell patterning is more precise using this latter approach. Each region is exposed to only one cell type, minimizing cross-contamination, and the interface is formed without relying on cell migration, ensuring sharp boundaries. See reference [2] and reference [3].
The method described in [2] utilizes a silicon substrate, which is not optically transparent. This makes the system incompatible with the inverted microscopes that are most widely employed in biology laboratories. On the other hand, although the method described in [3] employs transparent substrates, there is no firm locking mechanism to ensure accuracy in horizontal and vertical alignment.
Importantly, neither reference [2] nor reference [3] are easy to use or follow for those with standard training in cell biology. Furthermore, while some of the techniques are simpler, the quality of the patterned cell interface is not as good. Thus, there is a strong need for a low-cost and easy-to-use method to pattern a sharp interface between cell populations.
Any feature or combination of features described herein are included within the scope of the present invention provided that the features included in any such combination are not mutually inconsistent as will be apparent from the context, this specification, and the knowledge of one of ordinary skill in the art. Additional advantages and aspects of the present invention are apparent in the following detailed description and claims.